Improved Polymer Hemocompatibility for Blood-Contacting Applications via S-Nitrosoglutathione Impregnation.

Blood-contacting medical devices (BCMDs) are inevitably challenged by thrombi formation, leading to occlusion of flow and device failure. Ideal BCMDs seek to mimic the intrinsic antithrombotic properties of the human vasculature to locally prevent thrombotic complications, negating the need for systemic anticoagulation. An emerging category of BCMD technology utilizes nitric oxide (NO) as a hemocompatible agent, as the vasculature's endothelial layer naturally releases NO to inhibit platelet activation and consumption. In this paper, we report for the first time the novel impregnation of S-nitrosoglutathione (GSNO) into polymeric poly(vinyl chloride) (PVC) tubing via an optimized solvent-swelling method. Material testing revealed an optimized GSNO-PVC material that had adequate GSNO loading to achieve NO flux values within the physiological endothelial NO flux range for a 4 h period. Through in vitro hemocompatibility testing, the optimized material was deemed nonhemolytic (hemolytic index <2%) and capable of reducing platelet activation, suggesting that the material is suitable for contact with whole blood. Furthermore, an in vivo 4 h extracorporeal circulation (ECC) rabbit thrombogenicity model confirmed the blood biocompatibility of the optimized GSNO-PVC. Platelet count remained near 100% for the novel GSNO-impregnated PVC loops (1 h, 91.08 ± 6.27%; 2 h, 95.68 ± 0.61%; 3 h, 97.56 ± 8.59%; 4 h, 95.11 ± 8.30%). In contrast, unmodified PVC ECC loops occluded shortly after the 2 h time point and viable platelet counts quickly diminished (1 h, 85.67 ± 12.62%; 2 h, 54.46 ± 10.53%; 3 h, n/a; 4 h, n/a). The blood clots for GSNO-PVC loops (190.73 ± 72.46 mg) compared to those of unmodified PVC loops (866.50 ± 197.98 mg) were significantly smaller (p < 0.01). The results presented in this paper recommend further investigation in long-term animal models and suggest that GSNO-PVC has the potential to serve as an alternative to systemic anticoagulation in BCMD applications.

Bioinspired ultra-low fouling coatings on medical devices to prevent device-associated infections and thrombosis.

Addressing thrombosis and biofouling of indwelling medical devices within healthcare institutions is an ongoing problem. In this work, two types of ultra-low fouling surfaces (i.e., superhydrophobic and lubricant-infused slippery surfaces) were fabricated to enhance the biocompatibility of commercial medical grade silicone rubber (SR) tubes that are widely used in clinical care. The superhydrophobic (SH) coatings on the tubing substrates were successfully created by dip-coating in superhydrophobic paints consisting of polydimethylsiloxane (PDMS), perfluorosilane-coated hydrophobic zinc oxide (ZnO) and copper (Cu) nanoparticles (NPs) in tetrahydrofuran (THF). The SH surfaces were converted to lubricant-infused slippery (LIS) surfaces through the infusion of silicone oil. The anti-biofouling properties of the coatings were investigated by adsorption of platelets, whole blood coagulation, and biofilm formation in vitro. The results revealed that the LIS tubes possess superior resistance to clot formation and platelet adhesion than uncoated and SH tubes. In addition, bacterial adhesion was investigated over 7 days in a drip-flow bioreactor, where the SH-ZnO-Cu tube and its slippery counterpart significantly reduced bacterial adhesion and biofilm formation of Escherichia coli relative to control tubes (>5 log10 and >3 log10 reduction, respectively). The coatings also demonstrated good compatibility with fibroblast cells. Therefore, the proposed coatings may find potential applications in high-efficiency on-demand prevention of biofilm and thrombosis formation on medical devices to improve their biocompatibility and reduce the risk of complications from medical devices.

S-Nitroso-N-Acetyl-D-Penicillamine Modified Hyperbranched Polyamidoamine for High-Capacity Nitric Oxide Storage and Release.

Synthetic nitric oxide (NO)-donating materials have been shown to have many beneficial effects when incorporated into biomedical materials. When released in the correct dosage, NO has been shown to increase the biocompatibility of blood and tissue contacting materials, but materials are often limited in the amount of NO that can be administered over a period of time. To address this, hyperbranched polyamidoamine (HPAMAM) was modified with the S-nitrosothiol, S-nitroso-N-acetyl-D-penicillamine, and nitrosated to form a controlled, high-capacity NO-donating compound (SNAP-HPAMAM). This compound has the potential of modifying polymers to release NO over long periods of time by being blended into a variety of base polymers. Nitric oxide release was triggered by photoinitiation and through passive ion-mediated release seen under physiological conditions. A material that delivers the beneficial dose of NO over a long period of time would be able to greatly increase the biocompatibility of long-term implantable devices. Structural analysis of a generation 2 HPAMAM molecule was done through Fourier transform infrared spectroscopy (FTIR), 1H nuclear magnetic resonance spectroscopy (NMR), and matrix assisted laser desorption ionization, time of flight (MALDI-TOF) mass spectrometry. The NO capacity of the finalized generation 2 SNAP-HPAMAM compound was approximately 1.90 ± 0.116 µmol NO/mg. Quantification of the functional groups in the compound proved that an average of 6.40 ± 0.309 reactive primary amine sites were present compared to the 8 reactive sites on a perfectly synthesized generation 2 dendrimer. There is a substantial advantage of using the hyper-branched HPAMAM over purified dendrimers in terms of reduced labor and expense while still providing a high-capacity NO donor that can be blended into different polymer matrices.

Electrospun Bioabsorbable Fibers Containing S-Nitrosoglutathione for Tissue Engineering Applications.

Blended and coaxial fibers comprising polycaprolactone and gelatin, containing the endogenous nitric oxide (NO) donor S-nitrosoglutathione (GSNO), were electrospun. Both types of fibers had their NO release profiles tested under physiological conditions to examine their potential applications as biomedical scaffolds. The coaxial fibers exhibited a prolonged and consistent release of NO over the course of 4 d from the core-encapsulated GSNO, while the blended fibers had a large initial release and leaching of GSNO that was exhausted over a shorter period of time. Bacterial testing of both fiber scaffolds was conducted over a 24 h period against Staphylococcus aureus (S. aureus) and demonstrated a 3-log reduction in bacterial viability. In addition, no cytotoxic response was reported when the material was tested on mouse fibroblast cells in vitro. These fibrous matrices were also shown to support cell growth, attachment, and overall activity of fibroblasts when exposed to NO, especially when GSNO was encapsulated within coaxial fibers. From an application point of view, these NO-releasing fibers offer great potential in tissue engineering and biomedical applications because of the crucial role of NO in regulating a variety of biological processes in humans such as angiogenesis, tissue remodeling, and eliminating foreign pathogens.

Fabrication of Bacteria- and Blood-Repellent Superhydrophobic Polyurethane Sponge Materials.

Biofilm and thrombus formation on surfaces results in significant morbidity and mortality worldwide, which highlights the importance of the development of efficacious fouling-prevention approaches. In this work, novel highly robust and superhydrophobic coatings with outstanding multiliquid repellency, bactericidal performance, and extremely low bacterial and blood adhesion are fabricated by a simple two-step dip-coating method. The coatings are prepared combining 1H,1H,2H,2H-perfluorooctyltriethoxysilane (FAS-17)-coated hydrophobic zinc oxide and copper nanoparticles to construct hierarchical micro/nanostructures on commercial polyurethane (PU) sponges followed by polydimethylsiloxane (PDMS) treatment that is used to improve the binding degree between the nanoparticles and the sponge surface. The micro/nanotextured samples can repel various liquids including water, milk, coffee, juice, and blood. Relative to the original PU, the superhydrophobic characteristics of the fabricated sponge cause a significant reduction in the adhesion of bacteria (Staphylococcus aureus) by up to 99.9% over a 4-day period in a continuous drip-flow bioreactor. The sponge is also highly resistant to the adhesion of fibrinogen and activated platelets with ∼76 and 64% reduction, respectively, hence reducing the risk of blood coagulation and thrombus formation. More importantly, the sponge can sustain its superhydrophobicity even after being subjected to different types of harsh mechanical damage such as finger-wiping, knife-scratching, tape-peeling, hand-kneading, hand-rubbing, bending, compress-release (1000 cycles) tests, and 1000 cm sandpaper abrasion under 250 g of loading. Hence, this novel hybrid surface with robustness and the ability to resist blood adhesion and bacterial contamination makes it an attractive candidate for use in diverse application areas.

Nanoparticles Encapsulating Nitrosylated Maytansine To Enhance Radiation Therapy.

Radiotherapy remains a major treatment modality for cancer types such as non-small cell lung carcinoma (or NSCLC). To enhance treatment efficacy at a given radiation dose, radiosensitizers are often used during radiotherapy. Herein, we report a nanoparticle agent that can selectively sensitize cancer cells to radiotherapy. Specifically, we nitrosylated maytansinoid DM1 and then loaded the resulting prodrug, DM1-NO, onto poly(lactide-co-glycolic)-block-poly(ethylene glycol) (PLGA-b-PEG) nanoparticles. The toxicity of DM1 is suppressed by nanoparticle encapsulation and nitrosylation, allowing the drug to be delivered to tumors through the enhanced permeability and retention effect. Under irradiation to tumors, the oxidative stress is elevated, leading to the cleavage of the S-N bond and the release of DM1 and nitric oxide (NO). DM1 inhibits microtubule polymerization and enriches cells at the G2/M phase, which is more radiosensitive. NO under irradiation forms highly toxic radicals such as peroxynitrites, which also contribute to tumor suppression. The two components work synergistically to enhance radiotherapy outcomes, which was confirmed in vitro by clonogenic assays and in vivo with H1299 tumor-bearing mice. Our studies suggest the great promise of DM1-NO PLGA nanoparticles in enhancing radiotherapy against NSCLC and potentially other tumor types.

Zinc-oxide nanoparticles act catalytically and synergistically with nitric oxide donors to enhance antimicrobial efficacy.

The development of infection-resistant materials is of substantial importance as seen with an increase in antibiotic resistance. In this project, the nitric oxide (NO)-releasing polymer has an added topcoat of zinc oxide nanoparticle (ZnO-NP) to improve NO-release and match the endogenous NO flux (0.5-4 × 10-10 mol cm-2 min-1 ). The ZnO-NP is incorporated to act as a catalyst and provide the additional benefit of acting synergistically with NO as an antimicrobial agent. The ZnO-NP topcoat is applied on a polycarbonate-based polyurethane (CarboSil) that contains blended NO donor, S-nitroso-N-acetylpenicillamine (SNAP). This sample, SNAP-ZnO, continuously sustained NO release above 0.5 × 10-10 mol cm-2 min-1 for 14 days while samples containing only SNAP dropped below physiological levels within 24 h. The ZnO-NP topcoat improved NO release and reduced the amount of SNAP leached by 55% over a 7-day period. ICP-MS data observed negligible Zn ion release into the environment, suggesting longevity of the catalyst within the material. Compared to samples with no NO-release, the SNAP-ZnO films had a 99.03% killing efficacy against Staphylococcus aureus and 87.62% killing efficacy against Pseudomonas aeruginosa. A cell cytotoxicity study using mouse fibroblast 3T3 cells also noted no significant difference in viability between the controls and the SNAP-ZnO material, indicating no toxicity toward mammalian cells. The studies indicate that the synergy of combining a metal ion catalyst with a NO-releasing polymer significantly improved NO-release kinetics and antimicrobial activity for device coating applications. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 00A: 000-000, 2019.

Active Release of an Antimicrobial and Antiplatelet Agent from a Nonfouling Surface Modification.

Two major challenges faced by medical devices are thrombus formation and infection. In this work, surface-tethered nitric oxide (NO)-releasing molecules are presented as a solution to combat infection and thrombosis. These materials possess a robust NO release capacity lasting ca. 1 month while simultaneously improving the nonfouling nature of the material by preventing platelet, protein, and bacteria adhesion. NO's potent bactericidal function has been implemented by a facile surface covalent attachment method to fabricate a triple-action coating-surface-immobilized S-nitroso- N-acetylpenicillamine (SIM-S). Comparison of NO loading amongst the various branching configurations is shown through the NO release kinetics over time and the cumulative NO release. Biological characterization is performed using in vitro fibrinogen and Staphylococcus aureus assays. The material with the highest NO release, SIM-S2, is also able to reduce protein adhesion by 65.8 ± 8.9% when compared to unmodified silicone. SIM-S2 demonstrates a 99.99% (i.e., ∼4 log) reduction for S. aureus over 24 h. The various functionalized surfaces significantly reduce platelet adhesion in vitro, for both NO-releasing and non-NO-releasing surfaces (up to 89.1 ± 0.9%), demonstrating the nonfouling nature of the surface-immobilized functionalities. The ability of the SIM-S surfaces to retain antifouling properties despite gradual depletion of the bactericidal source, NO, demonstrates its potential use in long-term medical implants.

Antibacterial and Cellular Response Toward a Gasotransmitter-Based Hybrid Wound Dressing.

Biological processes such as infection, angiogenesis, and fibroblast proliferation and migration need to be regulated for effective healing of a wound. Failing to do so can delay the overall wound healing and add to the suffering and healthcare cost. Endogenous nitric oxide (NO) is a well-known gasotransmitter in the natural healing process in humans and other mammals. To utilize its inherent ability in the current study, an exogenous NO donor (S-nitroso-glutathione, GSNO) was integrated into a hybrid formulation consisting of a natural polymer (alginate) and a synthetic polymer (poly(vinyl alcohol) (PVA)). The alginate-PVA-GSNO dressings showed a sustained NO release for 72 h that resulted in 99.89 ± 0.40% and 98.93 ± 0.69% eradication of Staphylococcus aureus and Pseudomonas aeruginosa, respectively, which are among the most common causal agents of wound infections. The designed dressings resulted in a 3-fold increase in the proliferation of human endothelial cells when compared with control without GSNO showing its angiogenic potential. In addition, mouse fibroblast cells exposed to leachates from alginate-PVA-GSNO dressings showed significantly higher proliferation when compared to control alginate-PVA showing the NO release from exogenous GSNO in fibroblast proliferation. Fibroblast migration was shown to be much faster with GSNO-based dressings when compared to corresponding control dressings resulting in complete closure of an in vitro wound model within 48 h. The porous dressings also possessed important physical properties such as swelling, water vapor transmission, and moisture content that are desirable for effective wound healing. Overall, this study supports the possibility of using therapeutic alginate-PVA-GSNO dressing to provide a supportive environment for accelerated wound healing.

Multifunctional S-Nitroso-N-acetylpenicillamine-Incorporated Medical-Grade Polymer with Selenium Interface for Biomedical Applications.

Modern crises in implantable or indwelling blood-contacting medical devices are mainly due to the dual problems of infection and thrombogenicity. There is a paucity of biomaterials that can address both problems simultaneously through a singular platform. Taking cues from the body's own defense mechanism against infection and blood clotting (thrombosis) via the endogenous gasotransmitter nitric oxide (NO), both of these issues are addressed through the development of a layered S-nitroso-N-acetylpenicillamine (SNAP)-doped polymer with a blended selenium (Se)-polymer interface. The unique capability of the SNAP-Se-1 polymer composites to explicitly release NO from the SNAP reservoir as well as generate NO via the incorporated Se is reported for the first time. The NO release from the SNAP-doped polymer increased substantially in the presence of the Se interface. The Se interface was able to generate NO in the presence of S-nitrosoglutathione (GSNO) and glutathione (GSH), demonstrating the capability of generating NO from endogenous S-nitrosothiols (RSNO). Scanning electron microscopy-energy dispersive spectroscopy (SEM-EDS) traced distribution of elemental Se nanoparticles on the interface and the surface properties were evaluated by surface wettability and roughness. The SNAP-Se-1 efficiently inhibited the growth of bacteria and reduced platelet adhesion while showing minimal cytotoxicity, thus potentially eliminating the risks of systemic antibiotic and blood coagulation therapy. The SNAP-Se-1 exhibited antibacterial activity of ∼2.39 and ∼2.25 log reductions in the growth of clinically challenging adhered Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli. SNAP-Se-1 also significantly reduced platelet adhesion by 85.5% compared to corresponding controls. A WST-8-based cell viability test performed on NIH 3T3 mouse fibroblast cells provided supporting evidence for the potential biocompatibility of the material in vitro. These results highlight the prospective utility of SNAP-Se-1 as a blood-contacting infection-resistant biomaterial in vitro which can be further tuned by application specificity.

Synthesis and Characterization of Controlled Nitric Oxide Release from S-Nitroso-N-Acetyl-d-Penicillamine Covalently Linked to Polyvinyl Chloride (SNAP-PVC).

Polyvinyl chloride (PVC) is one of the most widely used polymers in medicine but has very poor biocompatibility when in contact with tissue or blood. To increase biocompatibility, controlled release of nitric oxide (NO) can be utilized to mitigate and reduce the inflammatory response. A synthetic route is described where PVC is aminated to a specified degree and then further modified by covalently linking S-nitroso-N-acetyl-d-penicillamine (SNAP) groups to the free primary amine sites to create a nitric oxide releasing polymer (SNAP-PVC). Controllable release of NO from SNAP-PVC is described using photoinitiation from light emitting diodes (LEDs). Ion-mediated NO release is also demonstrated as another pathway to provide a passive mechanism for NO delivery. The large range of NO fluxes obtained from the SNAP-PVC films indicate many potential uses in mediating unwanted inflammatory response in blood- and tissue-contacting devices and as a tool for delivering precise amounts of NO in vitro.

Achieving Long-Term Biocompatible Silicone via Covalently Immobilized S-Nitroso- N-acetylpenicillamine (SNAP) That Exhibits 4 Months of Sustained Nitric Oxide Release.

Ever since the role of endogenous nitric oxide (NO) in controlling a wide variety of biological functions was discovered approximately three decades back, multiple NO-releasing polymeric materials have been developed. However, most of these materials are typically short lived due to the inefficient incorporation of the NO donor molecules within the polymer matrix. In the present study, S-nitroso- N-acetyl penicillamine (SNAP) is covalently attached to poly(dimethylsiloxane) (PDMS) to create a highly stable nitric oxide (NO) releasing material for biomedical applications. By tethering SNAP to the cross-linker of PDMS, the NO donor is unable to leach into the surrounding environment. This is the first time that a sustainable NO release and bacterial inhibition for over 125 days has been achieved by any NO-releasing polymer with supporting evidence of potential long-term hemocompatibility and biocompatibility. The material proves to have very high antibacterial efficacy against Staphylococcus aureus by demonstrating a 99.99% reduction in the first 3 days in a continuous flow CDC bioreactor, whereas a similar inhibitory potential of 99.50% was maintained by the end of 1 month. Hemocompatibility of SNAP-PDMS was tested using a rabbit extracorporeal circuit (ECC) model over a 4 h period. Thrombus formation was greatly reduced within the SNAP-PDMS-coated ECCs compared to the control circuits, observing a 78% reduction in overall thrombus mass accumulation. These results demonstrate the potential of utilizing this material for blood and tissue contacting biomedical devices in long-term clinical applications where infection and unwanted clotting are major issues.

Biotemplated Synthesis and Characterization of Mesoporous Nitric Oxide-Releasing Diatomaceous Earth Silica Particles.

Diatomaceous earth (DE), a nanoporous silica material composed of fossilized unicellular marine algae, possesses unique mechanical, molecular transport, optical, and photonic properties exploited across an array of biomedical applications. The utility of DE in these applications stands to be enhanced through the incorporation of nitric oxide (NO) technology shown to modulate essential physiological processes. In this work, the preparation and characterization of a biotemplated diatomaceous earth-based nitric oxide delivery scaffold are described for the first time. Three aminosilanes [(3-aminopropyl)triethoxysilane (APTES), N-(6-aminohexyl)aminomethyltriethoxysilane (AHAMTES), and 3-aminopropyldimethylethoxysilane (APDMES)] were evaluated for their ability to maximize NO loading via the covalent attachment of N-acetyl-d-penicillamine (NAP) to diatomaceous earth. The use of APTES cross-linker resulted in maximal NAP tethering to the DE surface, and NAP-DE was converted to NO-releasing S-nitroso-N-acetyl-penicillamine (SNAP)-DE by nitrosation. The total NO loading of SNAP-DE was determined by chemiluminescence to be 0.0372 ± 0.00791 µmol/mg. Retention of diatomaceous earth's unique mesoporous morphology throughout the derivatization was confirmed by scanning electron microscopy. SNAP-DE exhibited 92.95% killing efficiency against Gram-positive bacteria Staphylococcus aureus as compared to the control. The WST-8-based cytotoxicity testing showed no negative impact on mouse fibroblast cells, demonstrating the biocompatible potential of SNAP-DE. The development of NO releasing diatomaceous earth presents a unique means of delivering tunable levels of NO to materials across the fields of polymer chemistry, tissue engineering, drug delivery, and wound healing.

Supplementation with l-carnitine does not reduce the efficacy of epirubicin treatment in breast cancer cells.

One of the cornerstones of therapy for invasive breast cancer includes the use of anthracyclines. Epirubicin, a stereoisomer of doxorubicin, is one of the commonly used anthracyclines. Anthracyclines while effective therapy for breast cancer, have their own unique toxicities, such as cardiomyopathy. l-Carnitine, a quarternary ammonium compound synthesized from methionine and lysine, is required for oxidative metabolism in mitochondria. Cardiac function is closely linked with oxidative metabolism whereby l-carnitine is an essential cofactor. A hypothesis is being investigated to determine if supplementation with carnitine in breast cancer patients treated with epirubicin will reduce the development of cardiac toxicity. We determined whether addition of l-carnitine altered the tumor cytotoxic effects of epirubicin using a number of in vitro cell viability assays in different breast cancer cell lines including BT549, MDA-MB-435, NCI-ADR-RES, MCF7 and T47D. Additionally we investigated the ability of cells to respond to l-carnitine following analysis of the expression of carnitine metabolic enzymes by RT-PCR. We determined that supplementation with l-carnitine had no effect on the ability of epirubicin to kill a variety of breast cancer cell lines. Additionally, no differences in the induction of apoptosis by epirubicin were observed. Furthermore, all cell lines examined expressed proteins required for carnitine uptake and use. Our data suggest that supplementation with l-carnitine does not impair the ability of epirubicin to kill breast cancer cells. These results suggest that supplementation with l-carnitine in patients undergoing epirubicin treatment could be safely used to reduce associated cardiotoxicities without fear that the efficacy of chemotherapy is jeopardized.

Tunable Nitric Oxide Release from S-Nitroso-N-acetylpenicillamine via Catalytic Copper Nanoparticles for Biomedical Applications.

The quest for novel therapies to prevent bacterial infections and blood clots (thrombosis) is of utmost importance in biomedical research due to the exponential growth in the cases of thrombosis and blood infections and the emergence of multi-drug-resistant strains of bacteria. Endogenous nitric oxide (NO) is a cellular signaling molecule that plays a pivotal role in host immunity against pathogens, prevention of clotting, and regulation of systemic blood pressure, among several other biological functions. The physiological effect of NO is dose dependent, which necessitates the study of its tunable release kinetics, which is the objective of this study. In the present study, polymer composites were fabricated by incorporating S-nitroso-N-acetylpenicillamine (SNAP) in a medical-grade polymer, Carbosil, and top-coated with varying concentrations of catalytic copper nanoparticles (Cu-NPs). The addition of the Cu-NPs increased the NO release, as well as the overall antimicrobial activity via the oligodynamic effect of Cu. SNAP (10 wt %) composites without Cu-NP coatings showed a NO flux of 1.32 ± 0.6 × 10-10 mol min-1 cm-2, whereas Cu-NP-incorporated SNAP films exhibited fluxes of 4.48 ± 0.5 × 10-10, 4.84 ± 0.3 × 10-10, and 11.7 ± 3.6 × 10-10 mol min-1 cm-2 with 1, 3, and 5 wt % Cu-NPs, respectively. This resulted in a significant reduction (up to 99.8%) in both gram-positive and gram-negative bacteria, with very low platelet adhesion (up to 92% lower) as compared to that of the corresponding controls. Copper leachates from the SNAP films were detected using the inductively coupled plasma-mass spectrometry technique and were found to be significantly lower in concentration than the recommended safety limit by the FDA. The cell viability test performed on mouse fibroblast 3T3 cells provided supportive evidence for the biocompatibility of the material in vitro.

A multi-defense strategy: Enhancing bactericidal activity of a medical grade polymer with a nitric oxide donor and surface-immobilized quaternary ammonium compound.

Although the use of biomedical devices in hospital-based care is inevitable, unfortunately, it is also one of the leading causes of the nosocomial infections, and thus demands development of novel antimicrobial materials for medical device fabrication. In the current study, a multi-defense mechanism against Gram-positive and Gram-negative bacteria is demonstrated by combining a nitric oxide (NO) releasing agent with a quaternary ammonium antimicrobial that can be covalently grafted to medical devices. Antibacterial polymeric composites were fabricated by incorporating an NO donor, S-nitroso-N-acetyl-penicillamine (SNAP) in CarboSil® polymer and top coated with surface immobilized benzophenone based quaternary ammonium antimicrobial (BPAM) small molecule. The results suggest that SNAP and BPAM individually have a different degree of toxicity towards Gram-positive and Gram-negative bacteria, while the SNAP-BPAM combination is effective in reducing both types of adhered viable bacteria equally well. SNAP-BPAM combinations reduced the adhered viable Pseudomonas aeruginosa by 99.0% and Staphylococcus aureus by 99.98% as compared to the control CarboSil films. Agar diffusion tests demonstrate that the diffusive nature of NO kills bacteria beyond the direct point of contact which the non-leaching BPAM cannot achieve alone. This is important for potential application in biofilm eradication. The live-dead bacteria staining shows that the SNAP-BPAM combination has more attached dead bacteria (than live) as compared to the controls. The SNAP-BPAM films have increased hydrophilicity and higher NO flux as compared to the SNAP films useful for preventing blood protein and bacterial adhesion. Overall the combination of SNAP and BPAM imparts different attributes to the polymeric composite that can be used in the fabrication of antimicrobial surfaces for various medical device applications. STATEMENT OF SIGNIFICANCE: A significant increase in the biomedical device related infections (BDRIs), inability of the currently existing antimicrobial strategies to combat them and a proportional rise in the associated morbidity demands development of novel antimicrobial surfaces. Some of the major challenges associated with the currently used therapeutics are: antibiotic resistance and cytotoxicity. In the current study, engineered polymeric composites with multi-defense mechanism were fabricated to kill bacteria via both active and passive mode. This was done by incorporating a nitric oxide (NO) donor S-nitroso-N-acetypenicillamine (SNAP) in a medical grade polymer (CarboSil®) and a benzophenone based quaternary ammonium antimicrobial small molecule (BPAM) was surface immobilized as the top layer. The developed biomaterial was tested with Gram-positive and Gram-negative strains and was found to be effective against both the strains resulting in up to 99.98% reduction in viable bacterial count. This preventative strategy can be used to fabricate implantable biomedical devices (such as catheters, stents, extracorporeal circuits) to not only significantly limit biofilm formation but also to reduce the antibiotic dose which are usually given post infections.

Corrosion Characteristics Dictate the Long-Term Inflammatory Profile of Degradable Zinc Arterial Implants.

There has been considerable recent interest to develop a feasible bioresorbable stent (BRS) metal. Although zinc and its alloys have many potential advantages, the inflammatory response has not been carefully examined. Using a modified wire implantation model, we characterize the inflammatory response elicited by zinc at high purity (4N) [99.99%], special high grade (SHG)[∼99.7%], and alloyed with 1 wt % (Zn-1Al), 3% (Zn-3Al), and 5.5% (Zn-5Al) aluminum. We found that inflammatory cells were able to penetrate the thick and porous corrosion layer that quickly formed around SHG, Zn-1Al, Zn-3Al, and Zn-5Al implants. In contrast, a delayed entrance of inflammatory cells into the corrosion layer around 4N zinc due to a significantly lower corrosion rate was associated with greater fibrous encapsulation, appearance of necrotic regions, and increased macrophage labeling. Interestingly, cell viability at the interface decreased from SHG, to Zn-1Al, and then Zn-3Al, a trend associated with an increased CD68 and CD11b labeling and capsule thickness. Potentially, the shift to intergranular corrosion due to the aluminum addition increased the activity of macrophages. We conclude that the ability of macrophages to penetrate and remain viable within the corrosion layer may be of fundamental importance for eliciting biocompatible inflammatory responses around corrodible metals.